Title: Functional Connectivity Changes during Early Recovery as a Marker for Relapse The relapsing nature of addiction is an obstacle to successful treatment. Future treatments need to integrate relevant brain biomarkers to guide individualized treatment for relapse prevention. We have cross-sectional data showing higher functional connectivity (FC) of nucleus accumbens (NAcc) and prefrontal regions in long- term abstinent substance users when compared to healthy controls, with intermediate FC in short-term abstinence. We found that lower FC in NAcc during short-term abstinence can predict relapse in the subsequent 6 months (75% accuracy). Cross-sectional results suggest that FC between NAcc and executive control regions may be a brain biomarker of vulnerability to relapse. In a pilot longitudinal FC study, we examined resting NAcc FC at 5 and 13 weeks of abstinence in individuals with stimulant use disorder (SUD), and collected relapse information 6 months later to define subsequent relapsers and abstainers. We found that NAcc FC (with frontal/parietal regions) decreased from 5 to 13 weeks of abstinence in subsequent relapsers, while it increased in subsequent abstainers. This time-varying factor may be a brain biomarker of relapse vulnerability. To disentangle this interaction effect, finer temporal resolution of both brin FC changes and relapse course is essential. In this project, we propose to collect data at 5, 9, 13 weeks during abstinence, with monthly follow-up interviews querying relapse status for 12 months. The overall objective of this proposal is to identify the dynamic changes in neural circuitry during the first 13 weeks of abstinence as potential brain biomarkers of relapse. Our central hypothesis is that those that subsequently relapse will initially have higher resting NAcc FC at 5 weeks followed by a reduction in FC at 13 weeks of abstinence and that this change can accurately predict relapse. Findings from this proposal will provide insight into the neurobiology of relapse vulnerability that will inform new treatment strategies needed to improve treatment outcome. Aim 1: To evaluate prediction accuracy of cross-sectional and longitudinal FC measures collected at 5, 9, and 13 weeks of abstinence. Hypothesis: Based on compensatory mechanisms hypothesis (SEC 3.1), we expect that subsequent abstainers will have stronger NAcc FC at each timepoint and a larger increase in NAcc FC across time when compared to subsequent relapsers and controls. Aim 2: To evaluate whether non-imaging measures add value to FC prediction models we will test prediction models using (i) only imaging variables, (ii) only non-imaging variables, and (iii) a combination of imaging and non-imaging variables as predictors of time to relapse. Hypothesis: Pilot data suggests that a combination of imaging and non-imaging variables with high temporal resolution will have high prediction accuracy (SEC 3.2). Exploratory Aim. To expand our examination of NAcc FC beyond resting state we will use a Stimulant-Cue Stroop task, which requires inhibitory control and reward processing. Hypothesis: Based on pilot data (SEC 3.4), subsequent abstainers will show a larger increase in task-related NAcc FC across time than subsequent relapsers and controls.